Mechanisms of drug action: Advanced mass spectrometry methods enable monitoring of tens of thousands of phosphorylation sites in proteins. This technology can potentially distinguish cellular signaling pathways that produce beneficial effects from those that produce unwanted side effects. Liuet al.treated mice with various agonists of the kappa opioid receptor (a G protein–coupled receptor) and monitored changes in phosphorylation over time in different brain regions. The phosphorylation patterns revealed distinct patterns of signaling in various brain tissues, some of which were associated with unwanted side effects.Science, this issue p.eaao4927

The A118G SNP (single nucleotide polymorphism) of the hMOPR [human MOPR (μ opioid receptor)] gene OPRM1 results in an amino acid substitution (N40D). Subjects homozygous for the 118G allele have been reported to require higher morphine doses to achieve adequate analgesia, and the 118G allele is more prevalent among drug abusers. However, changes in the MOPR protein associated with this SNP are unknown. Using a knockin mouse model (G/G mice; mice homozygous for the 112G allele of MOPR) that possesses the equivalent nucleotide/amino acid substitution (A112G; N38D) of the A118G SNP in the hMOPR gene, we investigated the N-linked glycosylation status of thalamic and striatal MOPR in G/G mice compared with A/A mice (wild-type mice homozygous for the 112A allele of MOPR). The molecular mass of MOPR determined by immunoblotting was lower in G/G mice than in A/A mice. Following treatment with peptide N-glycosidase F, which removes all N-linked glycans, both MOPR variants had an identical molecular mass, indicating that this discrepancy was due to a lower level of N-glycosylation of the MOPR in G/G mice. In Chinese-hamster ovary cells stably expressing hMOPRs, 118G/Asp⁴⁰-hMOPR had a lower molecular mass than 118A/Asn⁴⁰-hMOPR, which was similarly due to differential N-glycosylation. Pulse–chase studies revealed that the half-life of the mature form of 118G/Asp⁴⁰-hMOPR (~12 h) was shorter than that of 118A/Asn⁴⁰-hMOPR (~28 h). Thus the A118G SNP reduces MOPR N-glycosylation and protein stability.

Bioactivity-guided fractionation of Corydalis yanhusuo has resulted in the isolation of eight known isoquinoline alkaloids - tetrahydropalmatine, isocorypalmine, stylopine, corydaline, columbamine, coptisin, 13-methylpalmatine, and dehydrocorybulbine. The tertiary alkaloids were further analyzed by chiral HPLC to determine the ratios of d-and l-isomers. The isolated compounds were screened for their binding affinities at the dopamine D₁ receptor. Isocorypalmine had the highest affinity (K_i = 83 nM). The structure-affinity relationships of these alkaloids are discussed.

Salvinorin (Sal) A is a naturally occurring, selective κ opioid receptor (KOPR) agonist with a short duration of action in vivo. Pharmacological properties of a C(2) derivative, 2-methoxymethyl (MOM)-Sal B, were characterized. MOM-Sal B bound to KOPR with high selectivity and displayed ∼3-fold higher affinity than U50,488H [(trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide methanesulfonate] and Sal A. It acted as a full agonist at KOPR in guanosine 5′-O-(3-[³⁵S]thio)triphosphate binding and was ∼5- and ∼7-fold more potent than U50,488H and Sal A, respectively. In Chinese hamster ovary cells stably expressing KOPR, all three κ agonists internalized or down-regulated KOPR to similar extents, with MOM-Sal B being the most potent. In mice, MOM-Sal B (0.05–1 mg/kg s.c.) caused immediate and dose-dependent immobility lasting ∼3 h, which was blocked by norbinaltorphimine. In contrast, ambulation in a Y-maze was increased when rats received MOM-Sal B (1–5 mg/kg s.c.). In addition, MOM-Sal B (0.5–5 mg/kg i.p.) produced antinociception (hot-plate test) and hypothermia in a dose-dependent manner in rats. MOM-Sal B was more potent than U50,488H in both tests and more efficacious than U50,488H in the hot-plate test. These latter two in vivo effects were blocked by norbinaltorphimine, indicating KOPR-mediated actions. Sal A at 10 mg/kg elicited neither antinociception nor hypothermia 30 min after administration to rats. In summary, MOM-Sal B is a potent and efficacious KOPR agonist with longer lasting in vivo effects than Sal A.